Objective The aim of this study was to investigate the mechanism by which Beclin1 gene regulates rhabdomyosarcoma RD cells. Methods Beclin1 gene was overexpressed in RD cells using the transfection technique and was divided into experimental group and negative control group. At the mRNA and protein levels, RT-qPCR and western blotting were used to detect the expression of Beclin1 gene, respectively. Cell proliferation, migration, invasion and apoptosis were detected based on CCK-8 test, scratch healing test, Transwell test and TUNEL test. Results Beclin1 gene had a significantly greater mRNA expression in the experimental group (4.06±0.46) compared to the negative control group (P=0.008, P<0.01). The expression level of protein in the experimental group was (1.84±0.30), which was higher than that in the negative control group (P=0.032, P<0.05). Two group's growth curve were drawn based on its optical density value. The experimental group's level of cell growth was lower than that of the negative control group (P<0.05). Cell migration in the experimental group was noticeably worse than that in the negative control group (P<0.01), and the experimental group's capacity for invasion was observably weaker than that of the negative control group (P<0.01). But the rate of apoptosis was notably higher than that of the negative control group (P<0.01). Conclusion The overexpression of autophagy regulatory gene Beclin1 can significantly promote the apoptosis of RD cells and inhibit the proliferation, migration and invasion of RD cells.